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Sutter, G. & Moss, B. Nonreplicating vaccinia vector efficiently expresses recombinant genes. Proc. Natl Acad. Sci. USA 89, 10847-10851

Laboratory of Viral Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892.
Proceedings of the National Academy of Sciences (Impact Factor: 9.67). 12/1992; 89(22):10847-51. DOI: 10.1073/pnas.89.22.10847
Source: PubMed

ABSTRACT

Modified vaccinia Ankara (MVA), a highly attenuated vaccinia virus strain that has been safety tested in humans, was evaluated for use as an expression vector. MVA has multiple genomic deletions and is severely host cell restricted: it grows well in avian cells but is unable to multiply in human and most other mammalian cells tested. Nevertheless, we found that replication of viral DNA appeared normal and that both early and late viral proteins were synthesized in human cells. Proteolytic processing of viral structural proteins was inhibited, however, and only immature virus particles were detected by electron microscopy. We constructed an insertion plasmid with the Escherichia coli lacZ gene under the control of the vaccinia virus late promoter P11, flanked by sequences of MVA DNA, to allow homologous recombination at the site of a naturally occurring 3500-base-pair deletion within the MVA genome. MVA recombinants were isolated and propagated in permissive avian cells and shown to express the enzyme beta-galactosidase upon infection of nonpermissive human cells. The amount of enzyme made was similar to that produced by a recombinant of vaccinia virus strain Western Reserve, which also had the lacZ gene under control of the P11 promoter, but multiplied to high titers. Since recombinant gene expression is unimpaired in nonpermissive human cells, MVA may serve as a highly efficient and exceptionally safe vector.

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Available from: Gerd Sutter, Jun 12, 2015
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    • "Severely attenuated, transgenes inserted (Sutter and Moss, 1992; Kochneva et al., 2012) Various cancer model (Drexler et al., 1999; Carroll et al., 1997) Various tumors (Larocca and Schlom, 2011; Amato et al., 2012; Gómez et al., 2013) MVA-5T4*, MVAhup53 "
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    • "Dissemination within the host is precluded in most species, including humans, due to the extremely impaired ability to replicate in mammalian and, particularly, in human cells345. This results from a block in virion morphogenesis at a late stage of infection, thus, replication deficiency has no apparent effect on viral or recombinant gene expression345. MVA also showed an excellent safety record when administered during the smallpox eradication campaign in approximately 150,000 individuals, including many persons at risk for the conventional smallpox vaccines678. "
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    • "Clonal virus isolate F6 at passage 584 on primary chicken embryo fibroblasts (CEFs) was used for this study. Recombinant MVA constructs have been described previously and virus stocks were generated by standard methods (Sutter and Moss, 1992; Kremer et al., 2012). In brief, viruses were propagated on CEF, concentrated and purified by ultracentrifugation through sucrose, and titrated on CEF to determine infectious units (IU). "
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